The present invention relates to pharmaceutical compositions and methods for effecting neuronal activities, inhibiting angiogenesis and treating glutamate abnormalities, compulsive disorders, prostate diseases and cancers using pyrrolecarbonylimino derivatives as N-acetylated xcex1-linked Acidic Dipeptidase (NAALADase) enzyme inhibitors.
The NAALADase enzyme, also known as prostate specific membrane antigen (PSM or PSMA) and human glutamate carboxypeptidase II (GCP II), catalyzes the hydrolysis of the neuropeptide N-acetyl-aspartyl-glutamate (xe2x80x9cNAAGxe2x80x9d) to N-acetyl-aspartate (xe2x80x9cNAAxe2x80x9d) and glutamate. Based upon amino acid sequence homology, NAALADase has been assigned to the M28 family of peptidases. There is, as yet, no crystallographic evidence of the structure of the NAALADase enzyme.
Recent studies have implicated NAALADase in the pathogenesis of glutamate-mediated disorders. Neuropathological studies on post-mortem tissue from patients with amyotrophic lateral sclerosis (ALS) indicate large decreases of N-acetylaspartate (NAA) and N-acetylaspartylglutamate (NAAG) tissue concentrations occurring in association with neuronal degeneration, and increases of NAA and NAAG in cerebral spinal fluid (CSF) from patients with ALS. Concordantly, abnormal NAAG levels and NAALADase activity have also been observed in post-mortem prefrontal and limbic brain tissue of schizophrenic patients. Autopsy studies also suggest a strong correlation between NAAG/NAA and Alzheimer""s disease. In post-mortem brain tissue, NAA and NAAG levels were found to be selectively decreased in brain areas (hippocampus and amygdala) affected by Alzheimer""s disease pathology.
Glutamate serves as the predominant excitatory neurotransmitter in the central nervous system (CNS). Neurons release glutamate in greater quantities when they are deprived of oxygen, as may occur during an ischemic brain insult such as a stroke or a heart attack. This excess release of glutamate in turn causes over-stimulation (excitotoxicity) of N-methyl-D-aspartate (NMDA), AMPA, Kainate, MGR and MGluR receptors. When glutamate binds to these receptors, ion channels in the receptors open, permitting flows of ions across their cell membranes, e.g., Ca2+ and Na+ into the cells and K+ out of the cells. These flows of ions, especially the influx of Ca2+, cause over-stimulation of the neurons. The over-stimulated neurons secrete more glutamate, creating a feedback amplification effect which is believed to ultimately result in cell death via the production of proteases, lipases, and free radicals.
Excessive activation of glutamate receptors has been implicated in various neurological diseases and conditions (for example, spinal cord injury, epilepsy, stroke, Alzheimer""s disease, Parkinson""s Disease, Amyotrophic Lateral Sclerosis (ALS), Huntington""s Disease, schizophrenia, acute and chronic pain, ischemia and neuronal loss following hypoxia, hypoglycemia, ischemia, trauma, nervous insult, compulsive disorders (particularly drug and alcohol dependence), demyelinating diseases, peripheral neuropathies, and diabetic neuropathy), as well as in the generation of long-term potentiation, which is regarded as an electrophysiological manifestation of learning and memory. Specifically, the NMDA subtype of glutamate receptor appears to be involved in learning processes because the NMDA antagonist 2-amino-5-phosphonopentanoate (AP5) selectively impairs learning and blocks long-term potentiation in rats. It has thus been proposed that deterioration in glutamatergic systems accounts for impairment in cognitive function observed in aged animals or in Alzheimer""s disease.
In particular, glutamatergic abnormalities have been associated with schizophrenia. For example, phencyclidine (PCP) and other antagonists of N-methyl-D-aspartate (NMDA) receptors induce psychotomimetic properties in healthy individuals and exacerbate preexisting symptoms of schizophrenia, suggesting that a depression of glutamate transmission contributes to schizophrenia. Additionally, it has been reported that antagonists of non-NMDA receptors or pretreatments that attenuate glutamate release reduce mnemonic and other behavioral effects of NMDA receptor antagonists. Studies have also shown that stimulation of certain subtypes of mGlu receptors mediates presynaptic depression and decreases evoke release of glutamate.
Recent studies have also advanced a glutamatergic basis for compulsive disorders, particularly drug dependence. For example, neurophysiological and pathological effects of ethanol have been found to be mediated through the glutamatergic system. Specifically, acute exposure to ethanol disrupts glutamatergic neurotransmission by inhibiting ion flow through channels in glutamate receptors, whereas chronic exposure up-regulates the number of glutamate receptors and thereby increases ion flow. Acute withdrawal from ethanol results in hyperexcitability and seizures in the presence of up-regulated channels, thereby making postsynaptic neurons vulnerable to excitotoxic damage.
Post mortem examinations of histologically normal brains from alcoholics have shown that chronic alcoholism moderately increases the density of the NMDA subtype of glutamate receptors in the frontal cortex. This up-regulation may represent a stage of ethanol-induced chronic neurotoxicity. As such, neurobiological effects of alcoholism, including intoxication, withdrawal seizures, delirium tremens, Wernicke-Korsakoff syndrome and fetal alcohol syndrome, can be understood as a spectrum of the consequences of ethanol""s effect on the glutamatergic system. In this regard, alcoholism may be considered another member of the expanding family of glutamate-related neurological disorders.
The glutamatergic system has also been implicated in the behavioral effects of other abused drugs. For example, studies have shown that glutamatergic antagonists block motor-stimulating activities induced by amphetamine and cocaine, and glutamatergic agonists cause the same stereotypy as that produced by amphetamine. These results represent pharmacological evidence that the expression of the stereotypic effect of psychomotor stimulants involves the glutamatergic system.
Epidemiologic studies have revealed a strong correlation between drug dependence and other compulsive disorders. Additionally, a common genetic anomaly has been found among people with alcoholism, cocaine dependence, nicotine dependence, pathological gambling, attention deficit disorder (ADD), Tourette""s syndrome, compulsive overeating and obesity. Such disorders are believed to be manifestations of the effects of excitotoxicity.
Ischemic injury may occur as a focal or global disruption of blood supply. Following ischemic insult, widespread neuronal depolarization occurs. Depolarization stimulates release of the stored neurotransmitter glutamate and results in impaired capacity of glutamate uptake mechanisms. Impaired glutamate uptake and enhanced glutamate release contribute to sustained elevation of extracellular glutamate in ischemic tissue, and may result in tissue damage. As more damage occurs, more glutamate may be released. Although not limited to any particular theory, it is believed that by interfering with or eliminating this cascade of glutamate toxicity, the compositions and methods of the present convention may be clinically useful in curbing the progression of ischemic injury.
Afferent pain fibers of the A-xcex4 and C types have their primary cell bodies in the dorsal root ganglia; central extensions of these nerve cells project, via the dorsal root, to the dorsal horn of the spinal cord or to the nucleus of the trigeminal nerve; the peripheral terminations of these primary pain receptors are the branch nerve endings in the skin and other organs. Excitatory amino acids, including glutamate, and ATP are putative neurotransmitters at the dorsal horn terminus of primary A-xcex4 fibers. The conscious awareness or perception of pain occurs only when the pain impulses actually reach the thalamocortical level. Although not limited to any particular theory, it is believed that by interfering with such nerve impulses, the compositions and methods of the present invention will be clinically useful in limiting or eliminating pain.
Diabetic neuropathy is a slowly progressive, mixed sensorimotor and autonomic neuropathy. A variety of pathogenic mechanisms have been proposed for diabetic neuropathy, including alteration in nerve metabolism induced by ischemia and, in some cases, autoimmunity. Although not limited to any particular theory, it is believed that by interfering with or eliminating these effects, the compositions and methods of the present convention may be clinically useful in curbing the progression of diabetic neuropathy.
Excessive activation of glutamate receptors has been implicated in anxiety and anxiety disorders. Significantly higher glutamate plasma levels have been detected in patients with mood disorders than in comparison subjects. Studies also suggest that the pharmacological effect of anxiolytic agents is mediated through the glutamatergic system. Although not limited to any particular theory, it is believed that by interfering with or eliminating these effects, the compositions and methods of the present convention are expected to be clinically useful in curbing anxiolytic activity.
Based on the above findings, the present inventors tested and found NAALADase inhibitors to be efficacious in the pharmacotherapy of glutamate abnormalities, such as drug dependence, diabetic neuropathy, pain, schizophrenia, ischemic injury, and anxiety.
Most research and development activity to date has focused on blocking post-synaptic glutamate receptors with compounds such as NMDA antagonists, glycine antagonists, and other post-synaptic excitatory amino acid (EAA) receptor blockers. Unfortunately, these efforts have proven difficult because each receptor has multiple sites to which glutamate may bind; in addition, these agents produce severe toxicities even under normal conditions, thus limiting their clinical use. Although not limited to any one particular theory, it is believed that NAALADase inhibitors block glutamate release pre-synaptically without interacting with post-synaptic glutamate receptors. Since NAALADase inhibitors do not appear to alter basal glutamate levels, they may be devoid of the behavioral toxicities associated with post-synaptic glutamate antagonists.
In addition to glutamate, NAALADase has also been associated with prostate-specific membrane antigen (PSMA). In particular, it has been shown that PSMA cDNA confers NAALADase activity and that NAALADase and PSMA exhibit at least 86% homologous sequence identity. Carter et al., Proc. Natl. Acad. Sci., Vol. 93, pp. 749-753 (1996). The molecular cloning of PSMA has been reported as a potential prostate carcinoma marker and hypothesized to serve as a target for imaging and cytotoxic treatment modalities for prostate cancer. Additionally, PSMA antibodies, particularly indium-111 labelled and itrium labelled PSMA antibodies, have been described and examined clinically for the diagnosis and treatment of prostate cancer. PSMA is expressed in prostatic ductal epithelium and is present in seminal plasma, prostatic fluid and urine.
Applicants have found NAALADase inhibitors to be effective in treating prostate diseases, particularly prostate cancer. Although not limited to any particular theory, it is believed that NAALADase inhibitors inhibit PSMA activity. Since mAbs to PSMA have been found to target 23 non-prostate carcinomas (Lui et al., Science Research, Vol. 57, pp. 3629-34 (1997)), the present inventors hypothesize that NAALADase inhibitors would also be effective in treating non-prostate cancers, particularly in tissues where NAALADase resides, such as the brain, kidney and testis.
NAALADase has also been found in neovasculature (new blood vessels). The present inventors have discovered that NAALADase inhibitors inhibit or prevent growth of neovasculature (angiogenesis), thereby providing potential therapeutic applications in treating diseases dependent upon angiogenesis. Examples of angiogenesis-dependent diseases include without limitation rheumatoid arthritis, cardiovascular disease, neovascular diseases of the eye, peripheral vascular disorders, and dermatologic ulcers. Angiogenesis is also essential for normal physiological processes, such as growth, fertility and soft tissue wound healing.
Cancer is another disease dependent upon angiogenesis. Cancer tumor cells secrete or release angiogenic substances that activate nearby endothelial cells. These endothelial cells respond by expressing a cell autonomous pattern of behavior that culminates in the formation of new blood vessels. Since research has demonstrated that angiogenesis is necessary to sustain the growth, invasion and metastasis of cancer tumors, the neovasculature inhibiting activity of NAALADase inhibitors further supports their utility in treating all types of cancers.
In view of the broad range of potential applications, a need exists for new NAALADase inhibitors, pharmaceutical compositions comprising such inhibitors, and methods of their use.
The present invention relates to pharmaceutical compositions and methods of using pyrrolecarbonylimino derivatives to inhibit N-Acetylated xcex1-Linked Acidic Dipeptidase (NAALADase) enzyme activity, thereby effecting neuronal activities, inhibiting angiogenesis, and treating glutamate abnormalities, compulsive disorders, prostate diseases and cancers.
Specifically, the present invention relates to a method for inhibiting NAALADase enzyme activity in a mammal, comprising administering to said mammal an effective amount of a compound of formula I 
or a pharmaceutically acceptable equivalent, wherein:
m and n are independently 0, 1, 2 or 3;
Y is xe2x80x94R2 or xe2x80x94NHR2;
R1 and R2 are independently an aryl or heteroaryl group substituted with one or more substituent(s);
Z1 and Z2 are independently a moiety of formula II, III or IV 
X1 and X5 are independently CHR3 or NR3;
X2, X3, X4, X6, X7, X8 and X9 are independently CR3 or N; and
R3 is H or CH3.
The present invention further relates to a method for treating a glutamate abnormality in a mammal, comprising administering to said mammal an effective amount of a compound of formula I 
or a pharmaceutically acceptable equivalent, wherein:
m and n are independently 0, 1, 2 or 3;
Y is xe2x80x94R2 or xe2x80x94NHR2;
R1 and R2 are independently an aryl or heteroaryl group substituted with one or more substituent(s);
Z1 and Z2 are independently a moiety of formula II, III or IV 
X1 and X5 are independently CHR3 or NR3;
X2, X3, X4, X6, X7, X8 and X9 are independently CR3 or N; and
R3 is H or CH3;
provided that when m and n are independently 1, 2 or 3, Y is xe2x80x94NHR2, R2 is naphthyl substituted with 1, 2 or 3 sulfonic acid(s), Z1 and Z2 are each a moiety of formula II, X1 and X2 are each CH, X3 is NR3, and R3 is CH3, then the glutamate abnormality is not a demyelinating disease; and when the compound of formula I is suramin, then the glutamate abnormality is not a CNS neurodegenerative disorder or a demyelinating disease.
The present invention further relates to a method for effecting a neuronal activity in a mammal, comprising administering to said mammal an effective amount of a compound of formula I 
a pharmaceutically acceptable equivalent, wherein:
m and n are independently 0, 1, 2 or 3;
Y is xe2x80x94R2 or xe2x80x94NHR2;
R1 and R2 are independently an aryl or heteroaryl group substituted with one or more substituent(s);
Z1 and Z2 are independently a moiety of formula II, III or IV 
X1 and X5 are independently CHR3 or NR3;
X2, X3, X4, X6, X7, X8 and X9 are independently CR3 or N; and
R3 is H or CH3;
provided that when m and n are independently 1, 2 or 3, Y is xe2x80x94NHR2, R2 is naphthyl substituted with 1, 2 or 3 sulfonic acid(s), Z1 and Z2 are each a moiety of formula II, X1 and X2 are each CH, X3 is NR3, and R3 is CH3, then the effecting a neuronal activity is not treating a demyelinating disease; and when the compound of formula I is suramin, then the effecting a neuronal activity is not treating a CNS neurodegenerative disorder or a demyelinating disease.
The present invention further relates to a method for treating a prostate disease in a mammal, comprising administering to said mammal an effective amount of a compound of formula I 
or a pharmaceutically acceptable equivalent, wherein:
m and n are independently 0, 1, 2 or 3;
Y is xe2x80x94R2 or xe2x80x94NHR2;
R1 and R2 are independently an aryl or heteroaryl group substituted with one or more substituent(s);
Z1 and Z2 are independently a moiety of formula II, III or IV 
X1 and X5 are independently CHR3 or NR3;
X2, X3, X4, X6, X7, X8 and X9 are independently CR3 or N; and
R3 is H or CH3;
provided that when the compound of formula I is suramin, then the prostate disease is not prostate cancer.
The present invention further relates to a method for treating cancer in a mammal, comprising administering to said mammal an effective amount of a compound of formula I 
or a pharmaceutically acceptable equivalent, wherein:
m and n are independently 0, 1, 2 or 3;
Y is xe2x80x94R2 or xe2x80x94NHR2;
R1 and R2 are independently an aryl or heteroaryl group substituted with one or more substituent(s);
Z1 and Z2 are independently a moiety of formula II, III or IV 
X1 and X5 are independently CHR3 or NR3;
X2, X3, X4, X6, X7, X8 and X9 are independently CR3 or N; and
R3 is H or CH3;
provided that when the compound of formula I is suramin, then the cancer is not prostate cancer.
The present invention further relates to a method for inhibiting angiogenesis in a mammal comprising administering to said mammal an effective amount of a compound of formula I 
or a pharmaceutically acceptable equivalent, wherein:
m and n are independently 0, 1, 2 or 3;
Y is xe2x80x94R2 or xe2x80x94NHR2;
R1 and R2 are independently an aryl or heteroaryl group substituted with one or more substituent(s);
Z1 and Z2 are independently a moiety of formula II, III or IV 
X1 and X5 are independently CHR3 or NR3;
X2, X3, X4, X6, X7, X8 and X9 are independently CR3 or N; and R3 is H or CH3.
The present invention further relates to a pharmaceutical composition comprising:
(i) an effective amount of a compound of formula I for inhibiting NAALADase enzyme activity or angiogenesis, effecting a neuronal activity, or treating a glutamate abnormality, prostate disease or cancer 
xe2x80x83or a pharmaceutically acceptable equivalent, wherein:
m and n are independently 0, 1, 2 or 3;
Y is xe2x80x94R2 or xe2x80x94NHR2;
R1 and R2 are independently an aryl or heteroaryl group substituted with one or more substituent(s);
Z1 and Z2 are independently a moiety of formula II, III or IV 
X1 and X5 are independently CHR3 or NR3;
X2, X3, X4, X6, X7, X8 and X9 are independently CR3 or N; and
R3 is H or CH3;
provided that when m and n are independently 1, 2 or 3, Y is xe2x80x94NHR2, Z1 and Z2 are each a moiety of formula II, X1 and X2 are each CH, X3 is NR3, and R3 is CH3, then R2 is not naphthyl substituted with 1, 2 or 3 sulfonic acid(s); and provided that the compound of formula I is not suramin; and
(ii) a pharmaceutically acceptable carrier.
In addition to other references generally known in the art, applicants have previously disclosed data relating to the relationship between glutamate and various glutamate abnormalities, and the effectiveness of NAALADase inhibitors generally with regard to treating ischemic insult, diminished neurological function, alcohol dependence, nicotine dependence, cancer cell and tumor growth, angiogenesis, cocaine dependence, diabetes, pain, and hyperalgesia. Applicants hereby incorporate by reference, as though set forth herein in full, such figures and discussions from U.S. Pat. Nos. 5,672,592, 5,795,877, 5,863,536, 5,880,112 and 5,902,817, allowed U.S. patent applications Ser. Nos. 08/825,997, 08/833,628, 08/842,360 and 08/899,319 for which the issue fees have been paid, and International Publications Nos. WO 97/48399, WO 97/48400, WO 97/48409 and WO 98/53812. It would be expected that the compounds of the present invention would be effective as NAALADase inhibitors, and as such would have the same uses as the aforementioned compounds.
The compounds of the present invention are available from commercial suppliers or can be readily prepared by an ordinarily skilled artisan using standard techniques such as those disclosed in U.S. Pat. No. 5,859,046, herein incorporated by reference, as though set forth herein in full.